This R03 application focuses on the finding that deletion of a single ion channel gene in endothelial cells suppresses epileptic seizures in mice. As background, it is known that cerebral blood flow (CBF) initially increases in response to status epilepticus (SE) due to the high neuronal energy demand. But this adaptive increase in CBF dissipates during SE progression, and despite continued high neuronal metabolism, small cerebral arteries and arterioles begin to inappropriately constrict to limit CBF to the challenged neurons. This pathogenic vasoconstriction, termed the ?inverse hemodynamic response? (IHR), is thought to contribute to the progression of SE and its sequela of neuronal cell death. The mechanism of IHR is unknown. However, we have observed that Cre-mediated deletion of cation-conducting TRPC3 channels in endothelial cells prevents the IHR during SE and reduces susceptibility to pilocarpine-induced SE. In other words, deletion of a single TRPC channel isoform only in endothelial cells appears to exert a profound effect on neurovascular coupling and seizure susceptibility. This proposal seeks to generate an inducible and brain-specific endothelial TRPC3 knockout line and to use this novel mouse line to elucidate the role of TRPC3 channels expressed in cerebral vasculature endothelial cells in SE-induced IHR. Two aims will generate and characterize the inducible and brain-specific endothelial TRPC3 knockout line using the Slco1c1-CreERT2 driver line, and determine the role of brain endothelial cells in normal neurovascular coupling and SE-induced IHR. The studies rely on complementary areas of expertise pooled by a research team with expertise in epilepsy, vascular ion channels, and cerebrovascular reactivity. Techniques include immunohistochemistry, evaluation of changes in cerebral blood flow using laser speckle contrast imaging, and two-photon imaging of cerebral vasculatures. The findings of this project will yield a novel mouse line for the investigation of the functional role of cerebral vascular endothelial TRPC3 channels and will provide preliminary data for a future R01 project that will determine the underlying mechanisms of SE-induced IHR.